Q: Can you tell me what light and temperature conditions you keep the strain in? We are finding that we have to transfer the cultures every few days to keep them alive, which seems a bit strange - I can't imagine that you are continuously keeping a culture collection in exponential growth.
A: The strain is not in the perpetual collection. They were all grown at ~100uE. The cryo pull is split 1:1 with fresh medium based on the time frame established by the cryo test run. A subset is created on the day of shipment to cover 2 weeks in case of shipment failure. All of our perpetual cultures are kept in borosilicate test tubes with 18mL of media. The transfer schedule we use is anywhere from 1 week to 10 weeks depending on the culture. The inoculum can be anywhere from 1 drop to 3mL, again depending on the culture. Every strain is kept in triplicate;grandmother, mother and daughter. We set the inoculum so that there are still viable cells in the grandmother, not to be used but they are there in case of a catastrophic incubator failure, by the time we reinoculate a daughter set.